The investigation and/or analysis of ligand-molecule interactions and/or the electrochemical behaviour of biomolecules are fundamentally important in many fields, including biology, immunology, chemistry and pharmacology.
A number of analytical techniques can be used to investigate ligand-molecule interactions. For example, biological analytes can be detected or quantified based on ligand-specific binding between a ligand and a receptor. Common ligand/receptor binding pairs include antigen-antibody, hormone-receptor, drug-receptor, cell surface antigen-lectin, biotin-avidin, substrate-enzyme, and complementary nucleic acid strands. The analyte to be detected may be either member of the binding pair; alternatively, the analyte may be a ligand analogue that competes with the ligand for binding to the receptor.
Other analytical techniques utilise the oxidation or reduction of a molecule on the surface of a solid support. For example, glucose sensors may include an enzyme, such as glucose oxidase, which converts glucose into reaction products including hydrogen peroxide. A suitable electrode can then measure the formation of hydrogen peroxide as an electrical signal. The signal is produced following the transfer of electrons from the peroxide to the electrode, and under suitable conditions the enzyme catalysed flow of current is proportional to the glucose concentration in a sample. Alternatively, an electrode surface may be used in combination with current or impedance measuring elements for detecting a change in current or impedance in response to the presence of a ligand-receptor binding event.
Many of the aforementioned analytical techniques involve immobilising a molecule on a solid support. Other than immobilising the molecules, the solid support may play no role in subsequent chemical or biological investigations of the immobilised molecules. Alternatively, the solid support may interact with the immobilised molecule, such as when the solid support is an electrode which is used to investigate the electrochemistry of the molecule.
A common technique for the immobilization of molecules on the surface of a solid support is to covalently attach molecules onto a surface of a solid support that has previously been modified with an alkanethiol. Covalent attachment of the molecules to the surface prohibits diffusion of the molecules away from the solid support and also typically results in the formation of a film of molecules that is limited to a single monolayer, thereby limiting the required sample volume. The formation of self-assembled monolayers (“SAMs”) of molecules in this way has enabled the design of new interfaces for the study of ligand-molecule binding interactions as well as specific redox-active analytes. For example, monolayers have been formed via alkanethiol-gold linkage and related linkages between carboxylates and phosphonates and metal oxide surfaces. Monolayers formed on gold surfaces are particularly suited for studying biomolecular recognition at surfaces because the well-defined structures are amenable to detailed characterization at a molecular level by using, for example, scanning tunneling microscopy, atomic force microscopy as well as other optical and electrochemical bioanalytical techniques.
Solid supports having a monolayer of immobilised molecules are commonly referred to as “chips” or “sensor chips”. The sensor chips are routinely used in biosensor instruments where one or more properties of the immobilised molecules may be measured. A representative class of biosensor instrumentation is sold by Biacore AB (Uppsala, Sweden) under the trade name BIAcore™ (hereinafter referred to as “the BIAcore instrument”). The BIAcore instrument includes a light emitting diode, a sensor chip covered with a thin gold film, an integrated fluid cartridge and a photodetector. Molecules that are receptors of an analyte of interest are immobilised on the surface of the sensor chip and the chip is contacted with a flow of sample containing the analyte of interest. Any change in the surface optical characteristics of the sensor chip arising from the binding of the analyte of interest are then measured by detecting any intensity loss or “dip” in light that is reflected from the gold film on the surface of the sensor chip.
Numerous devices for determination of analytes that are based on the use of sensor chips are now available. However, many of the available sensor chips have some limitation with respect to sensitivity, test sample volume, reproducibility, speed of response, number of effective uses, or the range of detection. In the clinical setting, it is a goal to maximize the data obtainable from relatively small test sample volumes during analysis of fluids.
The discussion of the background to the invention herein is included to explain the context of the invention. This is not to be taken as an admission that any of the material referred to was published, known or part of the common general knowledge in any country. Further, throughout this specification reference may be made to documents for the purpose of describing various aspects of the invention. However, no admission is made that any document cited in this specification forms part of the common general knowledge in the art in any country.